Influence of receptor density on the patterns of b - adrenoceptor 2 desensitization

Sustained stimulation of the b -adrenoceptor leads to a desensitization of the receptor-mediated adenylyl cyclase stimulation. While 2 desensitization promoted by nanomolar concentrations of isoproterenol involves the phosphorylation of the b -adrenoceptor by protein 2 kinase A alone, both protein kinase Aand b-adrenoceptor kinase-mediated phosphorylation leading to the binding of b-arrestin contribute to the desensitization evoked by micromolar concentrations of agonist. In the present study, we assessed the influence of receptor density on the patterns of desensitization induced by these two different levels of stimulation. Murine L cells were transfected with a cDNA encoding the human b -adrenoceptor and clonal cell lines expressing various levels of b -adrenoceptor were used for the 2 2 Ž . study. In cell lines expressing the highest number of receptor, approx. 150 000 sitesrcell approx. 3000 fmolrmg of membrane proteins , pretreatment with micromolar concentrations of isoproterenol causes a desensitization pattern characterized by a reduction in both the potency and the efficacy of isoproterenol to further stimulate the adenylyl cyclase activity. In contrast, desensitization induced by 10 nM isoproterenol resulted only in a decrease in the potency of isoproterenol. This distinct pattern of desensitization is not seen in cells Ž . expressing 12 000 receptorsrcell approx. 200 fmolrmg of membrane proteins and, in that case, pretreatment with 10 nM isoproterenol leads to a reduction in both the sensitivity and the maximal response. Similar effects on the b-adrenoceptor-stimulated adenylyl cyclase were observed in these cells following treatment with dibutyryl cAMP. Receptor density therefore dramatically influences the pattern of desensitization evoked by low level of stimulation. The results also demonstrate that although different molecular events are involved in the desensitization evoked by different levels of stimulation, its phenotypic expression can be qualitatively identical in cells expressing a relatively small number of receptors. Hence, protein kinase A-mediated desensitization cannot be qualitatively distinguished from the b-adrenoceptor kinase-mediated process in these cells.